1. Field of the Invention
The present invention relates to aryl carbamate oligomers for making hydrolyzable prodrugs, oligomerized prodrugs including such oligomers, methods of synthesizing the aryl carbamate oligomers and oligomerized prodrugs, and methods of treatment employing the use of such polymeric prodrugs.
2. Related Art
Many peptides and proteins (collectively referred to herein as “polypeptides”) are potentially useful as therapeutic agents but lack an adequate means of administration. The usefulness of polypeptides as therapeutic agents is limited by the biological barriers that must be traversed before a polypeptide can reach its specific in vivo target. Parenterally administered polypeptides are readily metabolized by plasma proteases. Oral administration, which is perhaps the most attractive route of administration, is even more problematic. In the stomach, acid degrades and enzymes break down proteins. Those polypeptides that survive to enter the intestine intact are subjected to additional proteolysis as they are continuously barraged by a variety of enzymes, including gastric and pancreatic enzymes, exo- and endopeptidases, and brush border peptidases. As a result, passage of polypeptides from the lumen of the intestine into the bloodstream is severely limited. There is, therefore, a need in the art for means which enable parenteral and oral administration of therapeutic polypeptides.
Various strategies have been used in attempts to improve oral and parenteral delivery of polypeptides. Some of the approaches used include the use of enzyme inhibitors to slow the rate of degradation of proteins and peptides in the gastrointestinal tract; manipulation of pH to inactivate local digestive enzymes; use of permeation enhancers to improve the absorption of protein and peptides by increasing their paracellular and transcellular transports; use of nanoparticles as particulate carriers to facilitate intact absorption by the intestinal epithelium, especially, Peyer's patches, and to increase resistance to enzyme degradation; liquid emulsions to protect the drug from chemical and enzymatic breakdown in the intestinal lumen; and micelle formulations for poorly water solubulized drugs.
An important subset of the strategies for improving administration of polypeptides has been the conjugation of polypeptides to various moieties, such as polymeric moieties, to modify the physiochemical properties of polypeptide drugs to increase resistance to acidic and enzymatic degradation and to enhance penetration of such drugs across mucosal membranes. For example, Abuchowski and Davis have described various methods for derivatizating enzymes to provide water-soluble, non-immunogenic, in vivo stabilized products (“Soluble polymers-Enzyme adducts”, Enzymes as Drugs, Eds. Holcenberg and Roberts, J. Wiley and Sons, New York, N.Y., (1981)). Abuchowski and Davis discuss various ways of conjugating enzymes with polymeric materials, such as dextrans, polyvinyl pyrrolidones, glycopeptides, polyethylene glycol and polyamino acids. The resulting conjugated polypeptides are reported to retain their biological activities and solubility in water for parenteral applications. Furthermore, in U.S. Pat. No. 4,179,337, Davis et al. report that polypeptides, such as enzymes and insulin, can be coupled to polyethylene glycol or polypropropylene glycol having a molecular weight of 500 to 20,000 daltons to provide a physiologically active non-immunogenic water soluble polypeptide composition. The polyethylene glycol or polypropylene glycol is reported to protect the polypeptide from loss of activity and the composition can be injected into the mammalian circulatory system with substantially no immunogenic response. However, this approach is directed towards improving parenteral administration of polypeptides, not oral delivery.
Other researchers have shown that polyethylene glycol linked to a protein improves stability against denaturation and enzymatic digestion. (Boccu et al. Pharmacological Research Communication 14, 11-120 (1982)). However, these polymers do not contain components for enhancing membrane interaction. Thus, the resulting conjugates suffer from the same problems as noted above and are not suitable for oral administration.
Our own prior work involving the conjugation of polypeptides to amphiphilic oligomers, i.e., oligomers having hydrophilic and lipophilic characteristics, has been a major advance in the field of oral delivery of polypeptides. For example, U.S. Pat. No. 5,681,811 to Ekwuribe et al., and related U.S. Pat. Nos. 5,438,040 and 5,359,030, describe stabilized, conjugated polypeptide complexes including a therapeutic agent coupled to an oligomer that includes lipophilic and hydrophilic moieties. A preferred subset of the polypeptide-oligomer conjugates described in the '811 patent includes a polymer having a linear polyalkylene glycol moiety and a linear alkyl moiety.
U.S. Pat. No. 6,309,633 to Ekwuribe et al. describes a “partial prodrug” approach in which a polypeptide is conjugated to an oligomer having hydrophilic and lipophilic components, and the lipophilic component is hydrolyzable under physiological conditions. Conjugation of a polypeptide using the oligomers results in a “partial prodrug” in which the presence of the full oligomer assists in delivering the orally administered conjugate through the digestive tract into the bloodstream where the lipophilic component is hydrolyzed to leave the hydrophilic component (e.g., polyethylene glycol polymer). Hydrolysis of the lipophilic component in the bloodstream can render the conjugate bioactive (or improve bioactivity) and/or improve circulation half-life. However, there remains a significant need in the art for a means of amphiphilically conjugating polypeptides for oral delivery in a manner which does not eliminate useful bioactivity of the parent polypeptide.
Prodrug approaches are commonly used with small molecule therapeutics. For example, patented 6-MNA and paclitaxel prodrugs are known to make use of ester prodrugs of carboxylates and alcohols, while the APAZA™ compound (Nobex Corporation, Research Triangle Park, N.C.) is a prodrug in which two independently active small molecule drugs are bonded together by an azo linkage that is reductively cleaved in vivo (See U.S. Pat. Nos. 6,552,078, 6,541,508, 6,525,098, 6,436,990 and 6,380,405).
Garmen et al. describe a protein-PEG prodrug (Garman, A. J., and Kalindjian, S. B., FEBS Lett., 1987, 223, 361-365). The authors report the preparation of a maleic anhydride reagent from polydispersed MPEG5000 and conjugation to tissue plasminogen activator and urokinase. The reaction of amino acids with maleic anhydrides is commonly used in peptide sequencing chemistry. The hydrolysis of the maleyl-amide bond to reform the amine-containing drug is aided by the presence of the neighboring free carboxyl group and the geometry of attack set up by the double bond. Garman states that the native proteins of the maleic anhydride conjugates were released under physiological conditions, and administration via a prodrug increased clearance rates for the proteins by five to ten times.
More recently, in the development of a pegylated interferon, Roberts et al. have described the use of a degradable linkage between polydispersed PEG and interferon α-2b (Roberts, M. J. et al., Adv. Drug Delivery Rev., 2002, 54, 459-476). The authors reported that conjugating PEG to interferon at low pH (˜5) resulted in conjugates that were linked through a carbamate to one of the nitrogens of the imidazole ring of histidine. Over time, PEG was released from the protein. These PEG-interferon α-2b conjugates are known by the trade name of PEG-Intron® (Schering Corporation). While in vitro PEG-Intron® is reported to be more active than PEGASYS® (pegylated interferon with a non-hydrolyzable branched PEG, Hoffmann-La Roche), PEGASYS® is reported to be more efficacious in vivo.
Other efforts in the area of releasable PEG chemistry have focused on using 1, 6 or 1,4 benzyl elimination (BE) strategies (Lee, S., et al., Bioconjugate Chem., 2001, 12, 163-169; Greenwald, R. B. et al., U.S. Pat. No. 6,180,095, 2001; Greenwald, R. B., et al., J. Med. Chem., 1999, 42, 3657-3667.); the use of trimethyl lock lactonization (TML) (Greenwald, R. B. et al., J. Med. Chem., 2000, 43, 475-487); the coupling of PEG carboxylic acid to a hydroxy-terminated carboxylic acid linker (Roberts, M. J., J. Pharm. Sci., 1998, 87(11), 1440-1445), and PEG prodrugs involving families of MPEG phenyl ethers and MPEG benzamides linked to an amine-containing drug via an aryl carbamate (Roberts, M. J., et al., Adv. Drug Delivery Rev., 2002, 54, 459-476), including a prodrug structure involving a meta relationship between the carbamate and the PEG amide or ether (U.S. Pat. No. 6,413,507 to Bently et al.); and prodrugs involving a disulfide reduction mechanism as opposed to a hydrolysis mechanism (Zalipsky, S., et al., Bioconjugate Chem., 1999, 10(5), 703-707).
The present invention provides further developments regarding a prodrug approach in which a protein-oligomer prodrug is delivered through the digestive tract and into the bloodstream where the oligomer portion is released to yield the fully active protein.